Partial Folding and Unique ATP Function in Protein Phosphatase 2A Latency and Activation

Abstract

The function of metalloenzyme protein phosphatase 2A (PP2A) relies on proper formation of diverse heterotrimeric PP2A holoenzymes, comprising common scaffold and catalytic (PP2Ac) subunits and a variable regulatory subunit. Holoenzyme assembly is highly regulated by carboxylmethylation of PP2Ac. Our recent study demonstrated that the enzymatic reaction of PP2A methylation is highly responsive to PP2A activation via a direct binding of PP2A methyltransferase to the dynamic PP2A active site (Mol. Cell. 2011, 41(3):331-42). Interestingly, the entire protein fold of PP2Ac is highly dynamic and tends to partially unfold, which renders α4-binding that involves the inner structure of PP2Ac near the active site. α4-binding leads to an allosteric relay of conformational changes that perturbs the scaffold subunit binding site at the opposite surface. This unique mode of α4-binding underlies important mechanisms for stably latency of PP2Ac as well as dissociation of PP2A holoenzymes and recycling of PP2Ac in response to cell signaling. We further showed that PP2A phosphatase activator makes broad contacts with the structural elements surrounding the PP2A active site, which stabilizes the protein fold of apo-PP2Ac required for activation. PTPA-binding also defines a combined ATP-binding pocket that orients ATP phosphoryl groups to bind directly to the PP2A active site. This allows ATP to modulate the metal-binding preferences of the PP2A active site and drastically enhances binding of Mg2+ by 10,000-fold, which is crucial for acquisition of pSer/Thr-specific phosphatase activity. Consistent with the activation chaperone function of PTPA and activation-dependent PP2A methylation, PTPA stimulates both PP2A phosphatase activity and methylation. Collectively, our studies revealed a linear pathway for the biogenesis of PP2A holoenzymes, including stable latency, activation, methylation, and holoenzyme assembly, which is elegantly controlled by the protein dynamics and conformational switches of PP2Ac.